by High-Performance Liquid Chromatography

Abstract

Gossypol, a pigment in cottonseed, is a polyphe- nolic, binaphthyl dialdehyde. Due to steric hindrance between the functional groups of the molecule at the bond connecting the two naphthyl rings, gossypol exists as (+)- and (−)-isomers. Gossypol is physiologically active with the (−)-isomer appear- ing to be more active and causing temporary infertility in males. It is thus important to know the amounts of isomers in livestock feeds. A quantitative high-performance liquid chromatography (HPLC) procedure was developed for the separation of (+)- and (−)-gossypol contained in cottonseed. This method involves de- rivatization of gossypol with (R)-(−)-2-amino-1-propanol fol- lowed by HPLC separation employing either a Phenomenex Prodigy (5 µ, ODS-3, 100 × 3.2 mm) or a MetaChem Inertsil (5 µ, ODS-3, 100 × 3.0 mm) reversed-phase column eluted with 80% acetonitrile and 20% 10 mM KH2PO4 adjusted to pH 3.0 with H3PO4 at 1.0 mL/min. The (+)- and (−)-gossypol-2- amino- 1-propanol complexes eluted at roughly 1.4 and 2.6 min, re- spectively. It was found that gossypol from Upland (Gossypium hirsutum) seed was rich in the (+)-enantiomer, with the (+)- and (−)-enantiomers in a ratio of about 65:35, respectively, while gossypol from the seed of a Pima (G. barbadense) cultivar (S-6) was slightly richer in the (−)-enantiomer (46.8:53.2).